Evaluating the effectiveness of a structured, simulator-assisted, peer-led training on cardiovascular physical examination in third-year medical students: a prospective, randomized, controlled trial.

Background: Previous research suggests that cardiac examination skills in undergraduate medical students frequently need improvement. There are different ways to enhance physical examination (PE) skills such as simulator-based training or peer-assisted learning (PAL). Aim: The aim of this study was to evaluate the effectiveness of a structured, simulator-assisted, peer-led training on cardiovascular PE. Methods: Participants were third-year medical students at Leipzig University Faculty of Medicine. Students were randomly assigned to an intervention group (IG) and a control group (CG). In addition to standard curricular training, IG received a peer-led, simulator-based training in cardiac PE. Participant performance in cardiac PE was assessed using a standardized checklist with a maximum of 25 points. Primary outcome was assessed via checklist point distribution. Results: 89 students were randomised to either CG (n=43) or IG (n=46) with 70 completing the study. Overall, IG students performed significantly better than CG students did (max. points: 25, M±SD in IG was 17±3, in CG 12±4, p<.0001). Simple mistakes such as not using the stethoscope correctly were more frequent in CG students. Prior experience did not lead to a significant difference in performance. Conclusions: Structured, peer-led and simulator-assisted teaching sessions improve cardiac PE skills in this setting compared to control students that did not receive this training.

Oxidative stress stimulates α-tocopherol transfer protein in human trophoblast tumor cells BeWo.

α-Tocopherol transfer protein (α-TTP) has been identified as the major intracellular transport protein for the antioxidant vitamin E (α-tocopherol). Expression of α-TTP on the reproductive system has been described both in mouse uterus and lately in the human placenta. The aim of this study was to clarify if placental expression of α-TTP can be modified by substances causing oxidative reactions. The human choriocarcinoma cell line BeWo was, therefore, treated with two known pro-oxidants. α-TTP expression was determined with immunocytochemistry and evaluated by applying a semiquantitative score. The presence of pro-oxidants in BeWo cells induced α-TTP expression. We thus hypothesize that stimulation of α-TTP expression by oxidative stress, as this was induced by pro-oxidants, could be part of an antioxidant process occurring in the placenta in the aim of enhancing the supply of α-tocopherol. This process could occur both in normal pregnancies, as well as in pregnancy disorders presented with intensified oxidative stress. In that view, this model is proposed for further oxidative stress studies on trophoblast and placenta, on the grounds of clarifying the role of α-tocopherol in pregnancy physiology and pathophysiology.

A novel role for α-tocopherol transfer protein (α-TTP) in protecting against chloroquine toxicity.

Chloroquine (CQ) is a widely prescribed anti-malarial agent and is also prescribed to treat autoimmune diseases. Clinical treatment with CQ is often accompanied by serious side effects such as hepatitis and retinopathy. As a weak base, CQ accumulates in intracellular acidic organelles, raises the pH, and induces osmotic swelling and permeabilization of acidic organelles, which account for CQ-induced cytotoxicity. We reported previously that CQ treatment caused α-tocopherol transfer protein (α-TTP), a gene product of familial vitamin E deficiency, to change its location from the cytosol to the surface of acidic organelles. Here we show that α-TTP plays a novel role in protecting against CQ toxicity both in vitro and in vivo. In the presence of CQ, rat hepatoma McARH7777 cells, which do not express α-TTP endogenously, showed more severe cytotoxicity, such as larger vacuolation of acidic organelles and caspase activation, than α-TTP transfectant cells. Similarly, α-TTP knockout mice showed more severe CQ toxicity, such as hepatotoxicity and retinopathy, than wild-type mice. These effects were not ameliorated by vitamin E supplementation. In contrast to bafilomycin A1 treatment, which prevents CQ accumulation in cells by raising the pH of acidic organelles, α-TTP expression prevented CQ accumulation without affecting the pH of acidic organelles. Taken together, our data suggest that α-TTP protects against CQ toxicity by preventing CQ accumulation in acidic organelles through a mechanism distinct from vitamin E transport.

ATP-binding cassette transporter A1 is involved in hepatic alpha-tocopherol secretion.

Vitamin E (alpha-tocopherol) is an essential fat-soluble nutrient with antioxidant properties. alpha-Tocopherol transfer protein (alpha-TTP), the product of the gene responsible for familial isolated vitamin E deficiency, plays an important role in maintaining the plasma alpha-tocopherol level by mediating the secretion of alpha-tocopherol by the liver. However, the mechanisms underlying hepatic alpha-tocopherol secretion are not fully understood. This study was undertaken to elucidate the mechanism of alpha-tocopherol re-efflux from hepatocytes, the cells that have the most important role in regulating plasma-alpha-tocopherol concentrations. From in vitro experiments using [(3)H]alpha-tocopheryl acetate and McARH7777 cells that stably express alpha-tocopherol transfer protein (alpha-TTP), the following results were obtained. First, addition of apolipoprotein A-I (apoA-I), a direct acceptor of the ATP-binding cassette transporter A1 (ABCA1)-secreted lipids, increased alpha-tocopherol secretion in a dose-dependent manner. Second, probucol, an antiatherogenic compound reported to be an inactivator of ABCA1 reduced hepatic alpha-tocopherol secretion. Third, ABCA1-RNAi suppressed hepatic alpha-tocopherol secretion. In a mouse in vivo experiment, addition of 1% probucol to the diet decreased plasma alpha-tocopherol concentrations. These results strongly suggest that ABCA1 is substantially involved in hepatic alpha-tocopherol secretion.

Immunohistochemical localization of alpha-tocopherol transfer protein and lipoperoxidation products in human first-trimester and term placenta.

OBJECTIVE: Pregnancy is described as a state of oxidative stress arising from the high metabolic turnover taking place during feto-placental development and little is known about the balance of oxidation and antioxidation in early human pregnancy. The aim of this study was to analyze placental expression of alpha-tocopherol transfer protein (alpha-TTP) as the major transport protein for the antioxidant alpha-tocopherol as well as the placental expression of two lipoperoxidation products, malondialdehyde (MDA) and 4-hydroxy-2-nonenal (HNE) in early first-trimester and term human placenta. STUDY DESIGN: Placental tissue was obtained from 10 pregnancy interruptions at 6-8 weeks gestational age and 10 samples were obtained from term pregnancies after routine cesarean section. The placental expression of alpha-TTP, MDA and HNE has been investigated with immunohistochemistry by the use of specific human alpha-TTP, MDA and HNE antibodies. RESULTS: While MDA and HNE showed similar expression in first-trimester and term placenta, alpha-TTP expression was less in first-trimester syncytiotrophoblast as compared to term. In first-trimester specimen, alpha-TTP showed major expression in extravillous trophoblast. In amniotic epithelial cells, a rising tendency in all three parameters investigated from immature to mature cells could be documented. No direct correlation between alpha-TTP, MDA and HNE expression was detected. CONCLUSIONS: Our study shows the presence of alpha-TTP not only in term, but in first-trimester extravillous trophoblast, syncytiotrophoblast and amniotic epithelium. Furthermore, lipoperoxidation products MDA and HNE are also present in first-trimester and term placenta, documenting the presence of oxidative processes in the placenta from early on. It therefore seems possible that scavenging of reactive oxygen species (ROS) by alpha-tocopherol is already required in first-trimester human pregnancy, but the missing correlation to MDA and HNE expression leads to the speculation that alpha-TTP and its ligand alpha-tocopherol have functions beyond the antioxidative capacity of alpha-tocopherol in early pregnancy.

Influence of long-chain polyunsaturated fatty acid formula feeds on vitamin E status in preterm infants.

It has been recommended to supplement formulas for preterm infants with n-3 and n-6 long-chain polyunsaturated fatty acids (LCP) to improve growth, visual acuity, and neurodevelopmental performance. However, large amounts of LCP may increase lipid peroxidation and oxidative stress in preterm infants. We investigated if, under high supplementation of natural tocopherols, LCP addition to formula can be performed safely without causing tocopherol depletion in cell membranes. Thirty-one healthy preterm infants with gestational ages from 28 to 32 weeks were evaluated in a prospective, randomized study from birth to day 42. Nine infants received an n-3 and n-6 LCP-enriched formula (A), eleven infants a standard formula (B), and eleven infants breast milk (control group). Alpha- and gamma-tocopherol extracts were added to both formulas, amounting to five times the value in breast milk (2.3 mg/dL in both formulas versus 0.45 mg/dL in breast milk). Erythrocyte arachidonic acid (AA) and docosahexaenoic acid (DHA) in the phosphatidylethanolamine fraction were similar in the three groups over the study period, whereas a significant reduction of erythrocyte AA and DHA could be detected in the phosphatidylcholine fraction in all three groups from day 14 onwards, when compared to respective cord blood values, with lowest values in the standard formula group. Amazingly, levels of alpha- and gamma-tocopherol were higher in plasma, erythrocytes, platelets, monocytes, and polymorphonuclear leukocytes with LCP supplementation as compared to standard formula and breast milk from day 7 onwards, whereas in buccal mucosal cells, this was not the case until day 42. Gammatocopherol uptake in the LCP-supplemented group was also significantly higher in all cell fractions studied from day 7 onwards. We therefore hypothesize that the LCP supplementation used in formula A improves tocopherol solubility and stability in biological membranes. Under high-dose vitamin E addition to n-3 and n-6 LCP-supplemented formula, no evidence for tocopherol depletion and furthermore, high accumulation of tocopherols, can be detected in healthy preterm infants.

pH-dependent translocation of alpha-tocopherol transfer protein (alpha-TTP) between hepatic cytosol and late endosomes.

BACKGROUND: alpha-Tocopherol transfer protein (alpha-TTP), a member of the Sec14 protein family, plays an important role in transporting alpha-tocopherol, a major lipid-soluble anti-oxidant, in the cytosolic compartment of hepatocytes and is known as a product of the causative gene for familial isolated vitamin E deficiency. It has been shown that the secretion of hepatocyte alpha-tocopherol taken up with plasma lipoproteins is facilitated by alpha-TTP. To explore the mechanism of alpha-TTP mediated alpha-tocopherol secretion, we investigated drugs which may affect this secretion. RESULTS: We found that, in a hepatocyte cell culture system, intracellular alpha-tocopherol transport is impaired by chloroquine, an agent known for its function of elevating the pH in acidic compartments. Under chloroquine treatment, the diffuse cytosolic distribution of alpha-TTP changes to a punctate pattern. Double-staining experiments with endocytosis markers revealed that alpha-TTP accumulates transiently on the cytoplasmic surface of late endosomal membranes. This phenomenon is specific for hepatoma cell lines or primarily cultured hepatocytes. Other members of the Sec14 family, such as cellular retinaldehyde-binding protein (CRALBP) and supernatant protein factor (SPF), do not show this accumulation. Furthermore, we elucidate that the obligatory amino acid sequence for this function is located between amino acids 21 and 50, upstream of the N-terminal end of the lipid-binding domain. CONCLUSION: We hypothesize that a liver-specific target molecule for alpha-TTP exists on the late endosomal membrane surface. This transient binding may explain the mechanism of how alpha-tocopherol is transferred from late endosomes to cytosolic alpha-TTP.

Vitamin E and transfer proteins.

PURPOSE OF REVIEW: Recently, the intracellular transport as well as cellular uptake and excretion of alpha-tocopherol, the major representative of vitamin E, have been elucidated. RECENT FINDINGS: Alpha-tocopherol transfer protein has been identified as the major intracellular transport protein for vitamin E, mediating alpha-tocopherol secretion into the plasma via a non-Golgi-dependent pathway, while other binding proteins seem to play a less important role. New information has accumulated concerning the role of this protein in the transport and supply of vitamin E to tissues such as the central nervous system and the feto-maternal unit. The scavenger receptor class B type I receptor, a membrane-bound protein, is capable of transferring vitamin E into the cell, while the ATP-binding cassette transporter A1 can excrete vitamin E out of the cell. Advances in the area of vitamin E metabolism have shown that alpha-CEHC (2,5,7,8-tetramethyl-2-(2'-carboxyethyl)-6-hydroxychroman) and gamma-CEHC (2,7,8-trimethyl-2-(2'-carboxyethyl)-6-hydroxychroman) are formed by a cytochrome p450-mediated process, important for alpha and gamma-tocopherol excretion. SUMMARY: Insights into the regulation of vitamin E transport and metabolism on the cellular level have made enormous advances, showing the complex interplay of influx, trafficking, efflux and metabolism of this crucial antioxidant.